PROJECT SUMMARY/ABSTRACT The prevalence of childhood obesity has dramatically increased leading to the prediction of a rapid generational decline in life expectancy and to a surge in medical care costs. Offspring exposed to excessive gestational weight gain (GWG) have greater total fat mass (FM) and central adiposity in childhood and are more likely to be overweight or obese. Currently, 56% of pregnant women experience excessive GWG. In pregnancies that experience GWG, it is critical to understand factors that influence FM accumulation and direct adipose tissue (AT) distribution as these are important early drivers of obesity occurrence, disease risk, and severity of disease development. Polyunsaturated fatty acids (PUFA) influence adipocyte development and FM accrual in two ways: influencing cell differentiation and fatty acid metabolism regulation. Increased prenatal exposure to the n-3 fatty acid family, including docosahexaenoic acid (DHA), prevented adipocyte maturation and suppressed genes involved in lipogenesis and increased the expression of genes involved with ? oxidation. The overall net effect was a decrease in AT deposition. Observational studies associate greater maternal DHA levels with lower offspring FM, visceral AT and greater fat-free mass in childhood. However, data are lacking from RCT studying a contemporary U.S. based cohort where DHA is prenatally supplemented and offspring fat accrual and distribution are directly measured. This study will take advantage of an ongoing Phase III randomized, double blinded, and placebo controlled U.S. based pregnancy cohort (R01 HD083292, ClinicalTrials.gov ID: NCT02626299) in which n=400 women will be randomized to either 1000 mg/day or 200 mg/day of DHA during pregnancy. The purpose of the follow-up study is to determine how the prenatal dose of DHA interacts with GWG to influence offspring FM accrual. Data are lacking that examine important moderators and potential therapeutic targets that influence offspring growth and body composition changes during a critical period of the first 1,000 days suggested to be significant for programming the offspring phenotype. To develop effective interventions, it is important to understand nutrients that may protect against an offspring phenotype related to metabolic disease risk. Observational evidence in humans and basic animal research suggest that DHA, a nutrient generally low in the diet of US women, could be such a nutrient. This project is innovative, capitalizing on a large, ongoing, RCT of high and low-dose DHA supplementation of U.S. pregnant women, testing the efficacy of DHA supplementation in women with excessive GWG, and using direct measures of body composition (DXA). This data will inform the gap in knowledge regarding the impact of high levels of prenatal DHA supplementation on programming offspring FM accrual. The proposal will help identify the need for new recommendations for prenatal DHA supplementation by our National Academy of Medicine (DHA is not recognized in the US as an essential nutrient) and lead to evidence-based guidelines for DHA supplementation during pregnancy where none currently exist.